Cells of the renal medulla in vivo are exposed to extraordinarily high concentrations of the potent denaturant urea as a consequence of the renal concentrating mechanism. The molecular mechanism through which these cells respond to and tolerate this harsh environment has implications for the understanding of renal water and urea homeostasis, and for the understanding of and potential enhancement of resistance to metabolic stress in diverse pathophysiological contexts. The applicant has shown that renal epithelial cells uniquely and specifically exhibit a series of signaling responses to exogenous urea that has features of both mitogenic and stress signaling, and which includes activation of effectors of a receptor tyrosine kinase. The overarching hypothesis is that urea favorably regulates the balance between mitogenesis (i.e., cytoprotection) and apoptosis in a renal epithelial cell-specific fashion, and that this effect of urea is achieved indirectly through G-protein-coupled receptor-dependent transactivation of an EGF receptor family member via activation (ectodomain shedding) of heparin-binding epidermal growth factor. In Aim I, the mechanism through which urea increases D-cyclin expression in renal epithelial cells will be investigated as a correlate of urea-inducible pro-proliferative signaling, through a combination of pharmacological, biochemical, and molecular biological approaches. In Aim II, the mechanism through which urea protects from the pro-apoptotic effect of hypertonicity in renal epithelial cells will be investigated, with attention to the role of putative cytoprotective signaling intermediates previously shown to be activated by urea. In Aim III, in light of our newest preliminary data, the role of ectodomain shedding of heparin-binding epidermal growth factor (HB-EGF) will be assessed with respect to the acquisition of the urea-stressed molecular phenotype. In addition, the sufficiency of these and related signaling elements for urea signaling will be examined in models of heterologous expression.